This is a national stage of International Application Ser. No. PCT/EP97/04985, filed Sep. 11, 1997.
The present invention relates to metabotropic glutamate receptors (mGluRs). In particular, the invention relates to the mGlu7 receptor and novel applications thereof in models for neurological disease and as a target for neuroactive drugs.
L-glutamate is the major excitatory neurotransmitter in the central nervous system (CNS). Two major classes of glutamatergic receptors exist. The first class, the ionotropic receptors, which consists of NMDA, AMPA and kainate receptors, is responsible for fast synaptic transmission in the mammalian CNS. The second class, the metabotropic glutamate receptors (mGluR), exert actions on neurotransmission, synaptic plasticity and cellular excitation that are less well characterised.
Synaptic release of neurotransmitter in the nervous system is often influenced by presynaptic mGluRs which in turn respond to neurotransmitters released from the same nerve terminal or from terminals of other neurons. The mGluRs have diverse critical roles in forms of synaptic plasticity such as long term potentiation (LTP) or long term depression, forms of synaptic plasticity believed to be involved in learning and memory in vertebrates. Presynaptic mGluR autoreceptors respond to glutamate and influence the probability of neurotransmitter release from a nerve terminal. In general, the activation of presynaptic mGluRs (e.g. class III mGluRs activated by L-AP4), has been found to reduce transmitter release from synapses in many brain regions.
One member of the mGluR family, mGluR7, has been only poorly characterised to date. It is the most conserved member of the mGluR family, with only 6-8 amino acid differences observed between rat and human proteins. mGluR7 is widely distributed throughout the nervous system and is localised presynaptically close to neurotransmitter release sites. In the hippocampus, a high density of the mGlu7a isoform (note: to date only two isoforms of mGluR7, a and b, that differ only in their carboxy-terminal amino acid sequence have been characterized; Flor, P. J., van der Putten, H., Ruegg, D., Lukic, S., Leonhardt, T., Sansig, G., Knoepfel, T., and Kuhn, R. 1996. A novel splice variant of a metabotropic glutamate receptor, human mGluR7b; Neuropharmacology 36, 153-159) is found, in particular, in presynaptic terminals of excitatory cells that synapse on mGluR1xcex1 expressing GABA-ergic interneurons that also express somatostatin. Therefore, the input of this particular class of interneuron seems to be endowed by a particularly strong mGluR7-mediated autoregulation. Why this is so remains speculative for the moment and it has been suggested that this peculiar mGluR7-mediated autoregulation might relate to the role of the mGluR1xcex11-positive interneurons in the hippocampal network (Shigemoto et al., Nature (1996) 318:523-525). mGluR1xcex1+ cells receive glutamatergic input from axon collaterals of principal cells. The GABAergic terminals of mGluR1xcex1+ cells make synapses on principal cell distal dendrites, which also receive a direct excitatory input from the entorhinal cortex.
It has been hitherto generally accepted and shown that the glutamate analogue (S)-2-amino-4-phosphonobutanoate (L-AP4) induced presynaptic suppression of neurotransmitter release. In several instances, it has been proposed that such an effect could be mediated by activation of mGluR7 receptors. In other words, it has been proposed that activation of mGluR7 results in inhibition of neurotransmitter release. Building on this presumption, Shigemoto et al. (Op. Cit.) suggest that the high level of presynaptic mGluR7 may suppress the release of glutamate when action potentials arrive at high frequency, allowing glutamate release to follow only relatively low frequency synaptic firing. According to this hypothesis mGluR7 would work as a low pass filter and its gating function would allow low but not high frequency stimuli to be passed on effectively. This has implications for the use of glutamate agonists and/or antagonists in the therapy of disorders associated with synaptic plasticity, such as learning and memory disorders, epilepsy, pain and possibly ischaemia. According to the generally accepted model, a mGluR7 agonist would be effective in reducing transmitter release and a mGluR7 agonist would have more pronounced-effects on transmitter release in terminals that are particularly enriched in mGluR7 such as excitatory terminals onto mGluR1xcex1+ interneurons. Also, according to data and models proposed, so far, agonists of mGluR7 would have more pronounced inhibitory effects on excitatory terminals that release transmitter to interneurons. This would favour a decrease in inhibition exerted by the interneurons. Agonists of mGluR7 might therefore lower rather than enhance thresholds for seizures and epilepsy caused by decreased inhibition. Current models thus favour the development of antagonists rather than agonists of mGluR7 for the treatment of diseases such as epilepsy or relieving symptoms related to a reduction of neurotransmitter release.
We have used knockout mice in which the mGluR7 gene has been deleted in order to determine that this hypothesis and its implications are incorrect. mGluR7 facilitates rather than inhibiting glutamate release.
The present invention provides the use of a metabotropic glutamate receptor mGluR7 agonist for the facilitation of neurotransmitter release from a nerve ending. Moreover, the invention.provides transgenic knockout non-human mammals lacking the mGluR7 gene, suitable for studying mGluR7 and modulators thereof. Furthermore, the transgenic knockout mammals according to the invention are suitable for the study of epilepsy.